ABSTRACT
<p><b>OBJECTIVE</b>To investigate the mechanism that mda-7/IL-24 selectively kills hepatocellular carcinoma (HCC) HepG2 cells in vitro.</p><p><b>METHODS</b>HCC cell line HepG2 and normal liver cell line L02 were infected with Ad.mda-7. The expression of mda-7/IL-24 was detected by RT-PCR and ELISA, respectively. The apoptotic effects were confirmed by Hoechst staining and flow cytometry assay, respectively. Furthermore, Bcl-2 family proteins, cytochrome C, Smac/DIABLO and caspase-9 were determined by Western blot.</p><p><b>RESULTS</b>The exogenous mda-7/IL-24 gene was expressed in HepG2 and L02 cells infected with Ad.mda-7. Ad.mda-7 induced apoptosis in HepG2 but not in L02 cells in vitro. The induction of tumor cell apoptosis is correlated with the increasing expression of Bax and decreasing expression of Bcl-2 and Bcl-xL genes, then facilitated the releasing of cytochrome C and Smac/DIABLO from mitochondria to cytoplasm and increasing the expression of caspase-9, eventually, resulted in apoptosis.</p><p><b>CONCLUSION</b>Ad.mda-7 selectively induces growth inhibition and apoptosis in hepatocellular carcinoma HepG2 cells but not in normal L02 hepatocytes in vitro, and the mechanism might involve the decrease of Bcl-2 and Bcl-xL and increase of Bak expression, facilitating the release of cytochrome C and Smac/DIABLO from mitochondria in HCC cells.</p>
Subject(s)
Humans , Adenoviridae , Genetics , Apoptosis , Caspase 9 , Metabolism , Cytochromes c , Metabolism , Hep G2 Cells , Hepatocytes , Cell Biology , Interleukins , Genetics , Metabolism , Intracellular Signaling Peptides and Proteins , Metabolism , Mitochondria , Metabolism , Mitochondrial Proteins , Metabolism , Proto-Oncogene Proteins c-bcl-2 , Metabolism , Transfection , bcl-2-Associated X Protein , Metabolism , bcl-X Protein , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of melanoma differentiation associated gene-7/interleukin-24 (mda-7/IL-24) on the hepatocellular carcinoma cell lines and normal liver cell line in vitro.</p><p><b>METHODS</b>Hepatocellular carcinoma cell lines HepG2, SMMC7721, Hep3B, MHCC97L, M6 and normal liver cell line L02 were infected with Ad.mda-7. The gene expression of mda-7/IL-24 in these cell lines was confirmed by RT-PCR and ELISA assay. MTT assay and flow cytometry were used to study tumor cell proliferation and cell cycle in vitro. Hoechst staining and cytometry assay after Annexin-V and PI staining were studied to indicate the apoptosis effect.</p><p><b>RESULTS</b>It was confirmed by RT-PCR that the exogenous mda-7/IL-24 gene expressed in all of these cells. The mda-7/IL-24 protein product was confirmed by assaying the supernatant with ELISA. MTT and apoptosis test indicated mda-7/IL-24 can induce the hepatocellular carcinoma cell lines growth suppression, apoptosis in vitro but not in normal liver cell line L02, cell cycle test revealed mda-7/IL-24 can block cancer cell lines in G2/M but not in L02.</p><p><b>CONCLUSIONS</b>mda-7/IL-24 selectively induces growth suppression, apoptosis in hepatocellular carcinoma lines but not in normal liver cell in vitro.</p>
Subject(s)
Humans , Adenoviridae , Genetics , Apoptosis , Genetics , Physiology , Carcinoma, Hepatocellular , Genetics , Pathology , Cell Cycle , Genetics , Physiology , Cell Line , Cell Line, Tumor , Cell Proliferation , Genetic Vectors , Hepatocytes , Cell Biology , Interleukins , Genetics , Physiology , Liver Neoplasms , Genetics , Pathology , TransfectionABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of melanoma differentiation associated gene-7/interleukin 24 (MDA/IL-24) on human hepatocellular carcinoma cell lines HepG2, MHCC97L and Hep3B and normal liver cell line L02 with a different p53 state.</p><p><b>METHODS</b>The MDA-7/IL-24 gene was transfected into human hepatocellular carcinoma cell lines HepG2, MHCC97L and Hep3B and hepatocyte line L02 with a replication-incompetent adenovirus vector. The mRNA expression of MDA7/IL-24 in HepG2, MHCC97L, Hep3B and L02 cells was confirmed using RT-PCR. Protein expression was confirmed using ELISA assay. MTT assay and flow cytometry were used to study tumor cell proliferation and cell cycle in vitro. Hoechst and flow cytometry assay after annexin-V and PI staining were performed to indicate the apoptosis effect.</p><p><b>RESULTS</b>Exogenous MDA-7/IL-24 gene was expressed in HepG2, MHCC97L, Hep3B and L02 cells. The protein product of MDA-7/IL-24 was confirmed in the supernatant. MTT assay and apoptosis test indicated MDA-7/IL-24 could induce growth suppression and apoptosis of HepG2, MHCC97L and Hep3B but could not in L02. Cell cycle test revealed MDA-7/IL-24 could block those cancer cells in G2/M but not in the normal cell L02.</p><p><b>CONCLUSION</b>MDA-7/IL-24 selectively induces growth suppression and apoptosis in hepatocellular carcinoma lines HepG2, MHCC97L and Hep3B in vitro independent of the state of p53 gene but not in normal liver cell L02. This indicates MDA-7/IL-24 can be a perfect gene for gene therapy in hepatocellular carcinoma.</p>